Hantaviruses, members of the Bunyaviridae family are enveloped negative stranded emerging RNA viruses and category A pathogens that are transmitted to humans through aerosolized excreta of infected rodents. Hantaviruses have evolved a unique translation mechanism, operated by viral nucleocapsid protein (N), which preferentially facilitates the translation of viral mRNAs in host cells where cellular transcripts are competing for the same translation machinery. This translation mechanism is conceptually different from all translation mechanisms known to date, and facilitates the translation of capped mRNAs without the requirement of eIF4F cap binding complex. Hantaviral mRNAs contain a unique 42-52 nucleotide long 5' untranslated region (UTR) harboring a highly conserved triplet repeat sequence at the 5' terminus that specifically binds to the trimeric N protein. In addition, N also binds to the 40S ribosomal subunit via the ribosomal protein S19 (RPS19), located at the head region of 40S subunit. N preferentially loads 40S ribosomal subunits onto the viral mRNA 5' UTR and facilitates their translation in vitro. We will further characterize the role of 5' UTR in the translation of viral mRNAs by N-mediated translation mechanism in cells. We will determine whether hantaviral mRNAs specifically depend on N-mediated translation mechanism for their translation. We will also identify and characterize the RPS19 binding domain in N protein and demonstrate the role of N-RPS19 interaction in N-mediated translation strategy. We will determine whether N protein remains associated with the 40S subunit of 80S ribosome in polysome associated mRNAs. These studies will shed light on a novel mechanism of translation initiation that likely most negative stranded RNA viruses use to favor the translation of their mRNAs in host cells during infection. The proposed studies will also reveal novel targets for antiviral drug design in future. PUBLIC HEALTH RELEVANCE: There is no treatment for hantavirus associated disease at present. The major goal of this application is to delineate the mechanism of hantavirus replication in cells. In addition, the proposed studies will help in the identification of targets for antiviral drug design.